Vitamin E (tocopherol) is a fat-soluble vitamin found in many vegetable seed oils and leafy green vegetables. Vitamin E has many functions including acting as an antioxidant of lipids, protecting of cell membranes and prevention of damage to membrane associated enzymes. α-Tocopherol, often referred to as vitamin E, belongs to a class of lipid-soluble antioxidants that includes α, β, γ, and δ-tocopherols and α, β, γ, and δ-tocotrienols. Although α, β, γ, and δ-tocopherols and α, β, γ, and δ-tocotrienols are sometimes referred to collectively as “vitamin E,” vitamin E is more appropriately defined chemically as (α-tocopherol. α-Tocopherol is significant for human health, in part because it is readily absorbed and retained by the body, and therefore has a higher degree of bioactivity than other tocopherol species (Traber and Sies, Annu. Rev. Nutr. 16:321-347 (1996)). However, other tocopherols, such as β, γ, and δ-tocopherols, also have significant health and nutritional benefits.
Supplements may contain the alpha tocopherol that is either in the “d” form or a combination of the “d” and “l” forms. The “d” form is more active than the “l” form but when comparing supplements, an equivalent number of international units (IU) indicate equivalent activity. Less information is available about the action of the beta, gamma and delta tocopherols, but they appear to have different antioxidant effects. Vitamin E supplementation in humans may have a variety of beneficial effects including slowing the progression of Alzheimer's disease, preventing heart disease, improving immune function in the elderly, reducing the risk of cataracts and decreasing the pain associated with arthritis.
Photosynthetic bacteria and higher plants share a common set of enzymatic reactions for tocopherol synthesis, in which gamma-tocopherol methyltransferase (GMT) catalyzes the conversion of gamma-tocopherol to alpha-tocopherol in the final step of Vitamin E synthesis. The gene encoding GMT has been isolated and characterized from a variety of plant species including, pepper, soybean, Euglena, spinach and Arabidopsis. See, e.g., Shigeoka et al. (1992) Biochim Biophys Acta. 1128(2-3):220-6; GenBank Accession Nos. BM890961, AAD 38271, AF213481 and AF104220). In many plant oils (the main dietary source of tocopherols), alpha-tocopherol is typically present in small amounts while high levels of its biosynthetic precursor, gamma-tocopherol are generally present. Attempts to overexpress GMT in order to produce crops with higher alpha tocopherol content have shown that GMT overexpression can result in higher alpha tocopherol levels. See, e.g., Shintani et al. (1998) Science 282:2098-2100. However, such attempts have been hampered in view of the lack of efficient and stable methods of gene regulation in a variety of crops and plants.
Thus, there remains a need for compositions and methods for targeted regulation of the gamma-tocopherol methyltransferase (GMT) gene in plants to facilitate numerous applications such as, for example, the optimization of crop traits affecting nutritional value. In addition, such targeted regulation of GMT can be used to study biosynthetic pathways and gene function in plants.